Molecular genetic studies of control of mRNA stability in the gene for the key enzyme of methionine biosynthesis

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Expression of the gene for cystathionine gamma-synthase (CGS), the key enzyme of methionine biosynthesis in higher plants is feedback-regulated at the step of mRNA stability and the amino acid sequence in the exon 1 of CGS itself is involved in this regulation. In order to identity the factors that are involved in this regulation, a chimeric gene carrying green fluorescent protein gene (GFP) fused to the exon 1 of CGS gene and placed under the control of a cauliflower mosaic virus 35S promoter was constructed and introduced into wild-type Arabidopsis. The transgenic line termed GFPc4 was used to screen for those mutants that show higher GFP fluorescence relative to the autofluorescence of chloroplasts. Four of the mutants thus isolated seemed to affect the expression of CGS in traps. One of them showed elevated levels of both mRNAs for endogenous CGS and exogenous GFP fusion genes and reduced down-regulation of stability of mRNAs of both endogenous CGS and exogenous GFP fusion genes in response to methionine application. Transient expression analyses by electroporation to the mutant protoplasts were carried out A similar chimeric gene was constructed using E coli beta-glucuronidase gene as a reporter and the effects of methionine and S-adenosylmethionine (SAM) in the culture medium were tested. The results showed that in the mutant, the effects of both methionine and SAM were reduced as compared to the parental line. SAM has been shown to be the efftector of the post-transcriptional regulation of CGS expression. The results suggest that a mutant that affects the post-transcriptional regulation of CGS expression in trans.